Infant microbes and metabolites point to childhood neurodevelopmental disorders.

This study has followed a birth cohort for over 20 years to find factors associated with neurodevelopmental disorder (ND) diagnosis. Detailed, early-life longitudinal questionnaires captured infection and antibiotic events, stress, prenatal factors, family history, and more. Biomarkers including cord serum metabolome and lipidome, human leukocyte antigen (HLA) genotype, infant microbiota, and stool metabolome were assessed. Among the 16,440 Swedish children followed across time, 1,197 developed an ND. Significant associations emerged for future ND diagnosis in general and for specific ND subtypes, spanning intellectual disability, speech disorder, attention-deficit/hyperactivity disorder, and autism. This investigation revealed microbiome connections to future diagnosis as well as early emerging mood and gastrointestinal problems. The findings suggest links to immunodysregulation and metabolism, compounded by stress, early-life infection, and antibiotics. The convergence of infant biomarkers and risk factors in this prospective, longitudinal study on a large-scale population establishes a foundation for early-life prediction and intervention in neurodevelopment.

Microbial metabolite enhances immunotherapy efficacy by modulating T cell stemness in pan-cancer.

The immune checkpoint blockade (ICB) response in human cancers is closely linked to the gut microbiota. Here, we report that the abundance of commensal Lactobacillus johnsonii is positively correlated with the responsiveness of ICB. Supplementation with Lactobacillus johnsonii or tryptophan-derived metabolite indole-3-propionic acid (IPA) enhances the efficacy of CD8+ T cell-mediated αPD-1 immunotherapy. Mechanistically, Lactobacillus johnsonii collaborates with Clostridium sporogenes to produce IPA. IPA modulates the stemness program of CD8+ T cells and facilitates the generation of progenitor exhausted CD8+ T cells (Tpex) by increasing H3K27 acetylation at the super-enhancer region of Tcf7. IPA improves ICB responsiveness at the pan-cancer level, including melanoma, breast cancer, and colorectal cancer. Collectively, our findings identify a microbial metabolite-immune regulatory pathway and suggest a potential microbial-based adjuvant approach to improve the responsiveness of immunotherapy.

Dietary tryptophan metabolite released by intratumoral Lactobacillus reuteri facilitates immune checkpoint inhibitor treatment.

The use of probiotics by cancer patients is increasing, including among those undergoing immune checkpoint inhibitor (ICI) treatment. Here, we elucidate a critical microbial-host crosstalk between probiotic-released aryl hydrocarbon receptor (AhR) agonist indole-3-aldehyde (I3A) and CD8 T cells within the tumor microenvironment that potently enhances antitumor immunity and facilitates ICI in preclinical melanoma. Our study reveals that probiotic Lactobacillus reuteri (Lr) translocates to, colonizes, and persists within melanoma, where via its released dietary tryptophan catabolite I3A, it locally promotes interferon-γ-producing CD8 T cells, thereby bolstering ICI. Moreover, Lr-secreted I3A was both necessary and sufficient to drive antitumor immunity, and loss of AhR signaling within CD8 T cells abrogated Lr's antitumor effects. Further, a tryptophan-enriched diet potentiated both Lr- and ICI-induced antitumor immunity, dependent on CD8 T cell AhR signaling. Finally, we provide evidence for a potential role of I3A in promoting ICI efficacy and survival in advanced melanoma patients.

IL4I1 Is a Metabolic Immune Checkpoint that Activates the AHR and Promotes Tumor Progression.

Aryl hydrocarbon receptor (AHR) activation by tryptophan (Trp) catabolites enhances tumor malignancy and suppresses anti-tumor immunity. The context specificity of AHR target genes has so far impeded systematic investigation of AHR activity and its upstream enzymes across human cancers. A pan-tissue AHR signature, derived by natural language processing, revealed that across 32 tumor entities, interleukin-4-induced-1 (IL4I1) associates more frequently with AHR activity than IDO1 or TDO2, hitherto recognized as the main Trp-catabolic enzymes. IL4I1 activates the AHR through the generation of indole metabolites and kynurenic acid. It associates with reduced survival in glioma patients, promotes cancer cell motility, and suppresses adaptive immunity, thereby enhancing the progression of chronic lymphocytic leukemia (CLL) in mice. Immune checkpoint blockade (ICB) induces IDO1 and IL4I1. As IDO1 inhibitors do not block IL4I1, IL4I1 may explain the failure of clinical studies combining ICB with IDO1 inhibition. Taken together, IL4I1 blockade opens new avenues for cancer therapy.

Dietary and Microbial Oxazoles Induce Intestinal Inflammation by Modulating Aryl Hydrocarbon Receptor Responses.

Genome-wide association studies have identified risk loci associated with the development of inflammatory bowel disease, while epidemiological studies have emphasized that pathogenesis likely involves host interactions with environmental elements whose source and structure need to be defined. Here, we identify a class of compounds derived from dietary, microbial, and industrial sources that are characterized by the presence of a five-membered oxazole ring and induce CD1d-dependent intestinal inflammation. We observe that minimal oxazole structures modulate natural killer T cell-dependent inflammation by regulating lipid antigen presentation by CD1d on intestinal epithelial cells (IECs). CD1d-restricted production of interleukin 10 by IECs is limited through activity of the aryl hydrocarbon receptor (AhR) pathway in response to oxazole induction of tryptophan metabolites. As such, the depletion of the AhR in the intestinal epithelium abrogates oxazole-induced inflammation. In summary, we identify environmentally derived oxazoles as triggers of CD1d-dependent intestinal inflammatory responses that occur via activation of the AhR in the intestinal epithelium.

Indoleamine 2,3-dioxygenase 1 activation in mature cDC1 promotes tolerogenic education of inflammatory cDC2 via metabolic communication.

Conventional dendritic cells (cDCs), cDC1 and cDC2, act both to initiate immunity and maintain self-tolerance. The tryptophan metabolic enzyme indoleamine 2,3-dioxygenase 1 (IDO1) is used by cDCs in maintaining tolerance, but its role in different subsets remains unclear. At homeostasis, only mature CCR7+ cDC1 expressed IDO1 that was dependent on IRF8. Lipopolysaccharide treatment induced maturation and IDO1-dependent tolerogenic activity in isolated immature cDC1, but not isolated cDC2. However, both human and mouse cDC2 could induce IDO1 and acquire tolerogenic function when co-cultured with mature cDC1 through the action of cDC1-derived l-kynurenine. Accordingly, cDC1-specific inactivation of IDO1 in vivo exacerbated disease in experimental autoimmune encephalomyelitis. This study identifies a previously unrecognized metabolic communication in which IDO1-expressing cDC1 cells extend their immunoregulatory capacity to the cDC2 subset through their production of tryptophan metabolite l-kynurenine. This metabolic axis represents a potential therapeutic target in treating autoimmune demyelinating diseases.

Kynurenine importation by SLC7A11 propagates anti-ferroptotic signaling.

IDO1 oxidizes tryptophan (TRP) to generate kynurenine (KYN), the substrate for 1-carbon and NAD metabolism, and is implicated in pro-cancer pathophysiology and infection biology. However, the mechanistic relationships between IDO1 in amino acid depletion versus product generation have remained a longstanding mystery. We found an unrecognized link between IDO1 and cell survival mediated by KYN that serves as the source for molecules that inhibit ferroptotic cell death. We show that this effect requires KYN export from IDO1-expressing cells, which is then available for non-IDO1-expressing cells via SLC7A11, the central transporter involved in ferroptosis suppression. Whether inside the "producer" IDO1+ cell or the "receiver" cell, KYN is converted into downstream metabolites, suppressing ferroptosis by ROS scavenging and activating an NRF2-dependent, AHR-independent cell-protective pathway, including SLC7A11, propagating anti-ferroptotic signaling. IDO1, therefore, controls a multi-pronged protection pathway from ferroptotic cell death, underscoring the need to re-evaluate the use of IDO1 inhibitors in cancer treatment.

Interleukin-33 Induces the Enzyme Tryptophan Hydroxylase 1 to Promote Inflammatory Group 2 Innate Lymphoid Cell-Mediated Immunity.

Group 2 innate lymphoid cells (ILC2s) regulate immunity, inflammation, and tissue homeostasis. Two distinct subsets of ILC2s have been described: steady-state natural ILC2s and inflammatory ILC2s, which are elicited following helminth infection. However, how tissue-specific cues regulate these two subsets of ILC2s and their effector functions remains elusive. Here, we report that interleukin-33 (IL-33) promotes the generation of inflammatory ILC2s (ILC2INFLAM) via induction of the enzyme tryptophan hydroxylase 1 (Tph1). Tph1 expression was upregulated in ILC2s upon activation with IL-33 or following helminth infection in an IL-33-dependent manner. Conditional deletion of Tph1 in lymphocytes resulted in selective impairment of ILC2INFLAM responses and increased susceptibility to helminth infection. Further, RNA sequencing analysis revealed altered gene expression in Tph1 deficient ILC2s including inducible T cell co-stimulator (Icos). Collectively, these data reveal a previously unrecognized function for IL-33, Tph1, and ICOS in promoting inflammatory ILC2 responses and type 2 immunity at mucosal barriers.

Immune-regulated IDO1-dependent tryptophan metabolism is source of one-carbon units for pancreatic cancer and stellate cells.

Cancer cells adapt their metabolism to support elevated energetic and anabolic demands of proliferation. Folate-dependent one-carbon metabolism is a critical metabolic process underpinning cellular proliferation supplying carbons for the synthesis of nucleotides incorporated into DNA and RNA. Recent research has focused on the nutrients that supply one-carbons to the folate cycle, particularly serine. Tryptophan is a theoretical source of one-carbon units through metabolism by IDO1, an enzyme intensively investigated in the context of tumor immune evasion. Using in vitro and in vivo pancreatic cancer models, we show that IDO1 expression is highly context dependent, influenced by attachment-independent growth and the canonical activator IFNγ. In IDO1-expressing cancer cells, tryptophan is a bona fide one-carbon donor for purine nucleotide synthesis in vitro and in vivo. Furthermore, we show that cancer cells release tryptophan-derived formate, which can be used by pancreatic stellate cells to support purine nucleotide synthesis.

Essential Amino Acid Metabolites as Chemical Mediators of Host-Microbe Interaction in the Gut.

Amino acids are indispensable substrates for protein synthesis in all organisms and incorporated into diverse aspects of metabolic physiology and signaling. However, animals lack the ability to synthesize several of them and must acquire these essential amino acids from their diet or perhaps their associated microbial communities. The essential amino acids therefore occupy a unique position in the health of animals and their relationships with microbes. Here we review recent work connecting microbial production and metabolism of essential amino acids to host biology, and the reciprocal impacts of host metabolism of essential amino acids on their associated microbes. We focus on the roles of the branched-chain amino acids (valine, leucine, and isoleucine) and tryptophan on host-microbe communication in the intestine of humans and other vertebrates. We then conclude by highlighting research questions surrounding the less-understood aspects of microbial essential amino acid synthesis in animal hosts.

Intersection of the microbiome and immune metabolism in lupus.

Systemic lupus erythematosus is a complex autoimmune disease resulting from a dysregulation of the immune system that involves gut dysbiosis and an altered host cellular metabolism. This review highlights novel insights and expands on the interactions between the gut microbiome and the host immune metabolism in lupus. Pathobionts, invasive pathogens, and even commensal microbes, when in dysbiosis, can all trigger and modulate immune responses through metabolic reprogramming. Changes in the microbiota's global composition or individual taxa may trigger a cascade of metabolic changes in immune cells that may, in turn, reprogram their functions. Factors contributing to dysbiosis include changes in intestinal hypoxia, competition for glucose, and limited availability of essential nutrients, such as tryptophan and metal ions, all of which can be driven by host metabolism changes. Conversely, the accumulation of some host metabolites, such as itaconate, succinate, and free fatty acids, could further influence the microbial composition and immune responses. Overall, mounting evidence supports a bidirectional relationship between host immunometabolism and the microbiota in lupus pathogenesis.

Modulation of multiple sclerosis risk and pathogenesis by the gut microbiota: Complex interactions between host genetics, bacterial metabolism, and diet.

Multiple sclerosis (MS) is an autoimmune disease of the central nervous system, affecting nearly 2 million people worldwide. The etiology of MS is multifactorial: Approximately 30% of the MS risk is genetic, which implies that the remaining ~70% is environmental, with a number of factors proposed. One recently implicated risk factor for MS is the composition of the gut microbiome. Numerous case-control studies have identified changes in gut microbiota composition of people with MS (pwMS) compared with healthy control individuals, and more recent studies in animal models have begun to identify the causative microbes and underlying mechanisms. Here, we review some of these mechanisms, with a specific focus on the role of host genetic variation, dietary inputs, and gut microbial metabolism, with a particular emphasis on short-chain fatty acid and tryptophan metabolism. We put forward a model where, in an individual genetically susceptible to MS, the gut microbiota and diet can synergize as potent environmental modifiers of disease risk and possibly progression, with diet-dependent gut microbial metabolites serving as a key mechanism. We also propose that specific microbial taxa may have divergent effects in individuals carrying distinct variants of MS risk alleles or other polymorphisms, as a consequence of host gene-by-gut microbiota interactions. Finally, we also propose that the effects of specific microbial taxa, especially those that exert their effects through metabolites, are highly dependent on the host dietary intake. What emerges is a complex multifaceted interaction that has been challenging to disentangle in human studies, contributing to the divergence of findings across heterogeneous cohorts with differing geography, dietary preferences, and genetics. Nonetheless, this provides a complex and individualized, yet tractable, model of how the gut microbiota regulate susceptibility to MS, and potentially progression of this disease. Thus, we conclude that prophylactic or therapeutic modulation of the gut microbiome to prevent or treat MS will require a careful and personalized consideration of host genetics, baseline gut microbiota composition, and dietary inputs.

A tryptophan metabolite made by a gut microbiome eukaryote induces pro-inflammatory T cells.

The large intestine harbors microorganisms playing unique roles in host physiology. The beneficial or detrimental outcome of host-microbiome coexistence depends largely on the balance between regulators and responder intestinal CD4+ T cells. We found that ulcerative colitis-like changes in the large intestine after infection with the protist Blastocystis ST7 in a mouse model are associated with reduction of anti-inflammatory Treg cells and simultaneous expansion of pro-inflammatory Th17 responders. These alterations in CD4+ T cells depended on the tryptophan metabolite indole-3-acetaldehyde (I3AA) produced by this single-cell eukaryote. I3AA reduced the Treg subset in vivo and iTreg development in vitro by modifying their sensing of TGFß, concomitantly affecting recognition of self-flora antigens by conventional CD4+ T cells. Parasite-derived I3AA also induces over-exuberant TCR signaling, manifested by increased CD69 expression and downregulation of co-inhibitor PD-1. We have thus identified a new mechanism dictating CD4+ fate decisions. The findings thus shine a new light on the ability of the protist microbiome and tryptophan metabolites, derived from them or other sources, to modulate the adaptive immune compartment, particularly in the context of gut inflammatory disorders.

MYC promotes tryptophan uptake and metabolism by the kynurenine pathway in colon cancer.

Tumors display increased uptake and processing of nutrients to fulfill the demands of rapidly proliferating cancer cells. Seminal studies have shown that the proto-oncogene MYC promotes metabolic reprogramming by altering glutamine uptake and metabolism in cancer cells. How MYC regulates the metabolism of other amino acids in cancer is not fully understood. Using high-performance liquid chromatography (HPLC)-tandem mass spectrometry (LC-MS/MS), we found that MYC increased intracellular levels of tryptophan and tryptophan metabolites in the kynurenine pathway. MYC induced the expression of the tryptophan transporters SLC7A5 and SLC1A5 and the enzyme arylformamidase (AFMID), involved in the conversion of tryptophan into kynurenine. SLC7A5, SLC1A5, and AFMID were elevated in colon cancer cells and tissues, and kynurenine was significantly greater in tumor samples than in the respective adjacent normal tissue from patients with colon cancer. Compared with normal human colonic epithelial cells, colon cancer cells were more sensitive to the depletion of tryptophan. Blocking enzymes in the kynurenine pathway caused preferential death of established colon cancer cells and transformed colonic organoids. We found that only kynurenine and no other tryptophan metabolite promotes the nuclear translocation of the transcription factor aryl hydrocarbon receptor (AHR). Blocking the interaction between AHR and kynurenine with CH223191 reduced the proliferation of colon cancer cells. Therefore, we propose that limiting cellular kynurenine or its downstream targets could present a new strategy to reduce the proliferation of MYC-dependent cancer cells.

Distinct ecological fitness factors coordinated by a conserved Escherichia coli regulator during systemic bloodstream infection.

The ability of bacterial pathogens to adapt to host niches is driven by the carriage and regulation of genes that benefit pathogenic lifestyles. Genes that encode virulence or fitness-enhancing factors must be regulated in response to changing host environments to allow rapid response to challenges presented by the host. Furthermore, this process can be controlled by preexisting transcription factors (TFs) that acquire new roles in tailoring regulatory networks, specifically in pathogens. However, the mechanisms underlying this process are poorly understood. The highly conserved Escherichia coli TF YhaJ exhibits distinct genome-binding dynamics and transcriptome control in pathotypes that occupy different host niches, such as uropathogenic E. coli (UPEC). Here, we report that this important regulator is required for UPEC systemic survival during murine bloodstream infection (BSI). This advantage is gained through the coordinated regulation of a small regulon comprised of both virulence and metabolic genes. YhaJ coordinates activation of both Type 1 and F1C fimbriae, as well as biosynthesis of the amino acid tryptophan, by both direct and indirect mechanisms. Deletion of yhaJ or the individual genes under its control leads to attenuated survival during BSI. Furthermore, all three systems are up-regulated in response to signals derived from serum or systemic host tissue, but not urine, suggesting a niche-specific regulatory trigger that enhances UPEC fitness via pleiotropic mechanisms. Collectively, our results identify YhaJ as a pathotype-specific regulatory aide, enhancing the expression of key genes that are collectively required for UPEC bloodstream pathogenesis.

Tryptophan extends the life of cytochrome P450.

Powerfully oxidizing enzymes need protective mechanisms to prevent self-destruction. The flavocytochrome P450 BM3 from Priestia megaterium (P450BM3) is a self-sufficient monooxygenase that hydroxylates fatty acid substrates using O2 and NADPH as co-substrates. Hydroxylation of long-chain fatty acids (≥C14) is well coupled to O2 and NADPH consumption, but shorter chains (≤C12) are more poorly coupled. Hydroxylation of p-nitrophenoxydodecanoic acid by P450BM3 produces a spectrophotometrically detectable product wherein the coupling of NADPH consumption to product formation is just 10%. Moreover, the rate of NADPH consumption is 1.8 times that of O2 consumption, indicating that an oxidase uncoupling pathway is operative. Measurements of the total number of enzyme turnovers before inactivation (TTN) indicate that higher NADPH concentrations increase TTN. At lower NADPH levels, added ascorbate increases TTN, while a W96H mutation leads to a decrease. The W96 residue is about 7 Å from the P450BM3 heme and serves as a gateway residue in a tryptophan/tyrosine (W/Y) hole transport chain from the heme to a surface tyrosine residue. The data indicate that two oxidase pathways protect the enzyme from damage by intercepting the powerfully oxidizing enzyme intermediate (Compound I) and returning it to its resting state. At high NADPH concentrations, reducing equivalents from the flavoprotein are delivered to Compound I by the usual reductase pathway. When NADPH is not abundant, however, oxidizing equivalents from Compound I can traverse a W/Y chain, arriving at the enzyme surface where they are scavenged by reductants. Ubiquitous tryptophan/tyrosine chains in highly oxidizing enzymes likely perform similar protective functions.

Liquid-liquid phase separation of the prion protein is regulated by the octarepeat domain independently of histidines and copper.

Liquid-liquid phase separation (LLPS) of the mammalian prion protein is mainly driven by its intrinsically disordered N-terminal domain (N-PrP). However, the specific intermolecular interactions that promote LLPS remain largely unknown. Here, we used extensive mutagenesis and comparative analyses of evolutionarily distant PrP species to gain insight into the relationship between protein sequence and phase behavior. LLPS of mouse PrP is dependent on two polybasic motifs in N-PrP that are conserved in all tetrapods. A unique feature of mammalian N-PrP is the octarepeat domain with four histidines that mediate binding to copper ions. We now show that the octarepeat is critical for promoting LLPS and preventing the formation of PrP aggregates. Amphibian N-PrP, which contains the polybasic motifs but lacks a repeat domain and histidines, does not undergo LLPS and forms nondynamic protein assemblies indicative of aggregates. Insertion of the mouse octarepeat domain restored LLPS of amphibian N-PrP, supporting its essential role in regulating the phase transition of PrP. This activity of the octarepeat domain was neither dependent on the four highly conserved histidines nor on copper binding. Instead, the regularly spaced tryptophan residues were critical for regulating LLPS, presumably via cation-π interactions with the polybasic motifs. Our study reveals a novel role for the tryptophan residues in the octarepeat in controlling phase transition of PrP and indicates that the ability of mammalian PrP to undergo LLPS has evolved with the octarepeat in the intrinsically disordered domain but independently of the histidines.

Coupling and regulation mechanisms of the flavin-dependent halogenase PyrH observed by infrared difference spectroscopy.

Flavin-dependent halogenases are central enzymes in the production of halogenated secondary metabolites in various organisms and they constitute highly promising biocatalysts for regioselective halogenation. The mechanism of these monooxygenases includes formation of hypohalous acid from a reaction of fully reduced flavin with oxygen and halide. The hypohalous acid then diffuses via a tunnel to the substrate-binding site for halogenation of tryptophan and other substrates. Oxidized flavin needs to be reduced for regeneration of the enzyme, which can be performed in vitro by a photoreduction with blue light. Here, we employed this photoreduction to study characteristic structural changes associated with the transition from oxidized to fully reduced flavin in PyrH from Streptomyces rugosporus as a model for tryptophan-5-halogenases. The effect of the presence of bromide and chloride or the absence of any halides on the UV-vis spectrum of the enzyme demonstrated a halide-dependent structure of the flavin-binding pocket. Light-induced FTIR difference spectroscopy was applied and the signals assigned by selective isotope labeling of the protein moiety. The identified structural changes in α-helix and ß-sheet elements were strongly dependent on the presence of bromide, chloride, the substrate tryptophan, and the product 5-chloro-tryptophan, respectively. We identified a clear allosteric coupling in solution at ambient conditions between cofactor-binding site and substrate-binding site that is active in both directions, despite their separation by a tunnel. We suggest that this coupling constitutes a fine-tuned mechanism for the promotion of the enzymatic reaction of flavin-dependent halogenases in dependence of halide and substrate availability.

Hordeum vulgare CYP76M57 catalyzes C2 shortening of tryptophan side chain by C-N bond rearrangement in gramine biosynthesis.

The indole alkaloid gramine, 3-(dimethylaminomethyl)indole, is a defensive specialized metabolite found in some barley cultivars. In its biosynthetic process, the tryptophan (Trp) side chain is shortened by two carbon atoms to produce 3-(aminomethyl)indole (AMI), which is then methylated by N-methyltransferase (HvNMT) to produce gramine. Although side chain shortening is one of the crucial scaffold formation steps of alkaloids originating from aromatic amino acids, the gene and enzyme involved in the Trp-AMI conversion reactions are unknown. In this study, through RNA-seq analysis, 35 transcripts were shown to correlate with gramine production; among them, an uncharacterized cytochrome P450 (CYP) gene, CYP76M57, and HvNMT were identified as candidate genes for gramine production. Transgenic Arabidopsis thaliana and rice overexpressing CYP and HvNMT accumulate AMI, N-methyl-AMI, and gramine. CYP76M57, heterologously expressed in Pichia pastoris, was able to act on Trp to produce AMI. Furthermore, the amino group nitrogen of Trp was retained during the CYP76M57-catalyzed reaction, indicating that the C2 shortening of Trp proceeds with an unprecedented biosynthetic process, the removal of the carboxyl group and Cα and the rearrangement of the nitrogen atom to Cß. In some gramine-non-accumulating barley cultivars, arginine 104 in CYP76M57 is replaced by threonine, which abolished the catalytic activity of CYP76M57 to convert Trp into AMI. These results uncovered the missing committed enzyme of gramine biosynthesis in barley and contribute to the elucidation of the potential functions of CYPs in plants and undiscovered specialized pathways.

Slippy-Sloppy translation: a tale of programmed and induced-ribosomal frameshifting.

Programmed ribosomal frameshifting (PRF) is a key mechanism that viruses use to generate essential proteins for replication, and as a means of regulating gene expression. PRF generally involves recoding signals or frameshift stimulators to elevate the occurrence of frameshifting at shift-prone 'slippery' sequences. Given its essential role in viral replication, targeting PRF was envisioned as an attractive tool to block viral infection. However, in contrast to controlled-PRF mechanisms, recent studies have shown that ribosomes of many human cancer cell types are prone to frameshifting upon amino acid shortage; thus, these cells are deemed to be sloppy. The resulting products of a sloppy frameshift at the 'hungry' codons are aberrant proteins the degradation and display of which at the cell surface can trigger T cell activation. In this review, we address recent discoveries in ribosomal frameshifting and their functional consequences for the proteome in human cancer cells.